Sequencing of the human genome has provided an unparalleled opportunity to increase our knowledge of cellular function and disease to aid the discovery and development of more effective therapeutics. The availability of chemical probes that affect specific cellular proteins suspected to be involved in disease processes provides a critical tool for scientists to validate a molecular target for drug discovery. Chemical probes can also provide valuable specialty reagents for basic, translational and clinical research to establish a pharmacophore model or serve as cellular imaging tools or biomarkers. An integrated HTS approach involving a multidisciplinary group of scientists at Southern Research Institute is proposed for the NIH Roadmap Initiative to establish a state-of-art Molecular Library Screening Center. The approach involves an unique organizational plan to implement any cell, target, or organism-based assay up to BSL-3 from the scientific community, adapt it to a HTS format, optimize hits, and rapidly disseminate data to the PubChem database for public sector access. Five functional groups are proposed including an Assay Implementation, HTS Implementation, Synthetic Chemistry and Probe Development, Informatics, along with an Administrative and Management core group. The specific aims of this proposal are to 1) provide scientific expertise and laboratory resources to optimize and automate a diverse range of target-directed and phenotypic endpoints using biochemical, cell and organism-based assays which are applicable to HTS, 2) expand existing HTS and develop new high content screening (HCS) capacities to screen 100,000 compounds in up to 20 different assays on an annual basis, 3) develop second-generation chemical libraries to optimize leads with therapeutic potential, 4) develop a sophisticated database for distribution of screening results and structures of actives to PubChem, and 5) effectively manage a multidisciplinary group of scientists to insure innovative assay design and accomplishment of screening goals in a time and cost-efficient manner. The proposal includes the development of new screening tools involving HCS and organism-based screening assays through consultants and strategic alliances that tap additional scientific and engineering expertise outside the Institute. The over-arching goal of this program is to screen large chemical libraries in phenotypic or target-based assays and optimize molecules that can be used as probes to determine the function of newly identified cellular proteins and their role in normal and disease processes.